改变半抗原与载体蛋白的连接桥对提高水杨酸ELISA灵敏度的影.doc

改变半抗原与载体蛋白的连接桥对提高水杨酸ELISA灵敏度的影王树才　周　燮 中国图书分类号　R446.61 　　摘　要　目的：探讨水杨酸 SA 与载体蛋白间连接桥的改变对识别SA抗体的产生及SA ELISA灵敏度的影响。方法：将SA、4-氨基水杨酸、5-氨基水杨酸分别用不同的方法与载体蛋白相偶联，从而在SA苯环的C4、C5两个不同位点合成了3种免疫原：SA C5 -N N-p-AHA-BSA A 、SA C4 -N CH- CH2 3-CH N-BSA B 和SA C5 -NH-CH2-NH-BSA C 及4种包被物：SA C5 -N N-p-AHA-OVA、SA C4 -N CH- CH2 3-CH N-OVA、SA C5 -N CH- CH2 3-CH N-OVA、SA C5 -NH-CH2-NH-OVA。免疫新西兰白兔制备抗体，用ELISA法检测。结果：获得针对A和C的两种多克隆抗体，后一种能专一识别游离态水杨酸，改变包被物中连接桥的结构可提高SA ELISA的检测灵敏度，故选用SA C5 -N CH- CH2 3-CH N-OVA建立了SA ELISA。结论：SA与载体蛋白间连接桥的改变对识别SA抗体的产生和SA ELISA的灵敏度有很大影响。 　　关键词　水杨酸　半抗原载体连接桥　多克隆抗体　酶联免疫检测法 Effect of changing conjugation bridge between hapten and carrier protein on the sensitivity of salicylate elisa WANG Shu-Cai,ZHOU Xie. Department of Agronomy,Nanjing Agricultural University,Nanjing 210095 Abstract　Objective：To study the effect of changing conjugation bridge between hapten and carrier protein on the raising of antibodies against salicylic acid and the sensitivity of salicylate ELISA.Methods:Three kinds of salicylic acid SA immunogens:SA C5 -N N-p-AHA-BSA A 、SA C4 -N CH- CH2 3-CH N-BSA B and SA C5 -NH-CH2-NH-BSA C and four kinds of SA coating conjugates:SA C5 -N N-p-AHA-OVA、SA C4 -N CH- CH2 3-CH N-OVA、SA C5 -N CH- CH2 3-CH N-OVA and SA C5 -NH-CH2-NH-OVA were synthesized by linking SA、4-aminosalicylic acid and 5-aminosalicylic acid respectively to bovine serum albumin BSA and ovalbumin OVA through different conjugation methods.Raising of antibodies in rabbits and effect of changing conjugation bridge on the sensitivity of saliaylate ELISA were detected by ELISA.Results:Two kinds of polyclonal antibodies were raised in rabbits against immunogens A and C respectively.The antibodies against immunogen C possessed a high specificity .Using these antibodies and choosing SA C5 -N CH- CH2 3-CH N-OVA as a coating conjugate,a kind of ELISA was developed which displayed a linearity range from 0.064 to 200.000 nmol of SA.Conclusion:Conjngation bridge changing has a great effect on the raising o